Detergent solubilized HLA-A, B, DR, and other human major histocompatibility complex (MHC) antigens will be purified by affinity chromatography using specific cross-reactive xeno-antisera raised in rabbits. Purified antigens will be used to produce allospecific and locus-specific antibodies for serological and biochemical studies, in primates or by murine hybridomas in vitro. New human MHC and non-MHC differentiation antigens on T and B-lymphocyte subsets will be purified as they are defined and used to produce similar primate and hybridoma antibodies. Variant human MHC products expressed on T and B lymphoblastoid cell lines will be subjected to mutagenesis and immunoselection using specific alloantisera and hybridoma antibodies. Antigens, normal or mutated, will be biochemically and serologically characterized to determine the relative positions and interrelationships of antigenic determinants of single gene products. Human cytotoxic T-cell responses to the H-Y antigen and to lymphocytes infected with measles virus will be systematically examined to define in these systems the precise HLA specificities involved in self-restricted killing. The role of HLA antigens in macrophage presentation of antigens to helper T-cells will be studied. We will develop assays for human plaque forming cells. We will examine the distribution of MHC determinants on non-specific helper T-cells and the expression of MHC determinants on T-cell replacing factors. Immunogenicity and tolerogenicity of HLA antigens will be studied at the helper T-cell and B-cell level using in vitro assays for HLA-specific antibody responses. The precise conditions required for the binding of murine cytotoxic T-effector cells to lipid vesicles with membrane-integrated target cell membrane glycoproteins will be determined. The role of H-2D and H-2K molecules and other MHC products in promoting specific binding will be investigated. The importance of the molecular orientation of target proteins and the composition and fluidity of the membrane will be examined. Effectors will be conjugated to antigen-loaded lipid vesicles with entrapped 6-carboxyfluorescein to isolate a purified effector population using a fluorescence-activated cell sorter.